Tyrosine kinase inhibitors

ABSTRACT

The present invention relates to pyrazolo-pyrimidinyl compounds which inhibit, regulate and/or modulate tyrosine kinase signal transduction, compositions which contain these compounds, and methods of using them to treat tyrosine kinase-dependent diseases and conditions, such as angiogenesis, cancer, tumor growth, atherosclerosis, age related macular degeneration, diabetic retinopathy, inflammatory diseases, and the like in mammals.

RELATED APPLICATIONS

This application claims the benefit under 35 U.S.C. §119(e) of U.S.Provisional Application 60/123,902, filed Mar. 11, 1999.

BACKGROUND OF THE INVENTION

The present invention relates to compounds which inhibit, regulateand/or modulate tyrosine kinase signal transduction, compositions whichcontain these compounds, and methods of using them to treat tyrosinekinase-dependent diseases and conditions, such as angiogenesis, cancer,tumor growth, atherosclerosis, age related macular degeneration,diabetic retinopathy, inflammatory diseases, and the like in mammals.

Tyrosine kinases are a class of enzymes that catalyze the transfer ofthe terminal phosphate of adenosine triphosphate to tyrosine residues inprotein substrates. Tyrosine kinases are believed, by way of substratephosphorylation, to play critical roles in signal transduction for anumber of cell functions. Though the exact mechanisms of signaltransduction is still unclear, tyrosine kinases have been shown to beimportant contributing factors in cell proliferation, carcinogenesis andcell differentiation.

Tyrosine kinases can be categorized as receptor type or non-receptortype. Receptor type tyrosine kinases have an extracellular, atransmembrane, and an intracellular portion, while non-receptor typetyrosine kinases are wholly intracellular.

The receptor-type tyrosine kinases are comprised of a large number oftransmembrane receptors with diverse biological activity. In fact, abouttwenty different subfamilies of receptor-type tyrosine kinases have beenidentified. One tyrosine kinase subfamily, designated the HER subfamily,is comprised of EGFR, HER2, HER3, and HER4. Ligands of this subfamily ofreceptors include epithileal growth factor, TGF-α, amphiregulin, HB-EGF,betacellulin and heregulin. Another subfamily of these receptor-typetyrosine kinases is the insulin subfamily, which includes INS-R, IGF-IR,and IR-R. The PDGF subfamily includes the PDGF-α and β receptors. CSFIR,c-kit and FLK-II. Then there is the FLK family which is comprised of thekinase insert domain receptor (KDR), fetal liver kinase-1 (FLK-1), fetalliver kinase-4 (FLK-4) and the fms-like tyrosine kinase-1 (flt-1). ThePDGF and FLK families are usually considered together due to thesimilarities of the two groups. For a detailed discussion of thereceptor-type tyrosine kinases, see Plowman et al., DN&P 7(6):334-339,1994, which is hereby incorporated by reference.

The non-receptor type of tyrosine kinases is also comprised of numeroussubfamilies, including Src, Frk, Btk, Csk, Abl, Zap70, Fes/Fps, Fak,Jak, Ack, and LIMK. Each of these subfamilies is further sub-dividedinto varying receptors. For example, the Src subfamily is one of thelargest and includes Src, Yes, Fyn, Lyn, Lck, Blk, Hck, Fgr, and Yrk.The Src subfamily of enzymes has been linked to oncogenesis. For a moredetailed discussion of the non-receptor type of tyrosine kinases, seeBolen Oncogene, 8:2025-2031 (1993), which is hereby incorporated byreference.

Both receptor-type and non-receptor type tyrosine kinases are implicatedin cellular signaling pathways leading to numerous pathogenicconditions, including cancer, psoriasis and hyperimmune responses.

Several receptor-type tyrosine kinases, and the growth factors that bindthereto, have been suggested to play a role in angiogenesis, althoughsome may promote angiogenesis indirectly (Mustonen and Alitalo, J. CellBiol. 129:895-898, 1995). One such receptor-type tyrsoine kinase isfetal liver kinase 1 or FLK-1. The human analog of FLK-1 is the kinaseinsert domain-containing receptor KDR, which is also known as vascularendothelial cell growth factor receptor 2 or VEGFR-2, since it bindsVEGF with high affinity. Finally, the murine version of this receptorhas also been called NYK (Oelrichs et al., Oncogene 8(1):11-15, 1993).VEGF and KDR are a ligand-receptor pair that play an important role inthe proliferation of vascular endothelial cells, and the formation andsprouting of blood vessels, termed vasculogenesis and angiogenesis,respectively.

Angiogenesis is characterized by excessive activity of vascularendothelial growth factor (VEGF). VEGF is actually comprised of a familyof ligands (Klagsburn and D'Amore, Cytokine & Growth Factor Reviews7:259-270, 1996). VEGF binds the high affinity membrane-spanningtyrosine kinase receptor KDR and the related fms-like tyrosine kinase-1,also known as Flt-1 or vascular endothelial cell growth factor receptor1 (VEGFR-1). Cell culture and gene knockout experiments indicate thateach receptor contributes to different aspects of angiogenesis. KDRmediates the mitogenic function of VEGF whereas Flt-1 appears tomodulate non-mitogenic functions such as those associated with cellularadhesion. Inhibiting KDR thus modulates the level of mitogenic VEGFactivity. In fact, tumor growth has been shown to be susceptible to theantiangiogenic effects of VEGF receptor antagonists. (Kim et al., Nature362, pp. 841-844, 1993).

Solid tumors can therefore be treated by tyrosine kinase inhibitorssince these tumors depend on angiogenesis for the formation of the bloodvessels necessary to support their growth. These solid tumors includehistiocytic lymphoma, cancers of the brain, genitourinary tract,lymphatic system, stomach, larynx and lung, including lungadenocarcinoma and small cell lung cancer. Additional examples includecancers in which overexpression or activation of Raf-activatingoncogenes (e.g., K-ras, erb-B) is observed. Such cancers includepancreatic and breast carcinoma. Accordingly, inhibitors of thesetyrosine kinases are useful for the prevention and treatment ofproliferative diseases dependent on these enzymes.

The angiogenic activity of VEGF is not limited to tumors. VEGF accountsfor most of the angiogenic activity produced in or near the retina indiabetic retinopathy. This vascular growth in the retina leads to visualdegeneration culminating in blindness. Ocular VEGF mRNA and protein areelevated by conditions such as retinal vein occlusion in primates anddecreased pO2 levels in mice that lead to neovascularization.Intraocular injections of anti-VEGF monoclonal antibodies or VEGFreceptor immunofusions inhibit ocular neovascularization in both primateand rodent models. Regardless of the cause of induction of VEGF in humandiabetic retinopathy, inhibition of ocular VEGF is useful in treatingthe disease.

Expression of VEGF is also significantly increased in hypoxic regions ofanimal and human tumors adjacent to areas of necrosis. VEGF is alsoupregulated by the expression of the oncogenes ras, raf, src and mutantp53 (all of which are relevant to targeting cancer). Monoclonalanti-VEGF antibodies inhibit the growth of human tumors in nude mice.Although these same tumor cells continue to express VEGF in culture, theantibodies do not diminish their mitotic rate. Thus tumor-derived VEGFdoes not function as an autocrine mitogenic factor. Therefore, VEGFcontributes to tumor growth in vivo by promoting angiogenesis throughits paracrine vascular endothelial cell chemotactic and mitogenicactivities. These monoclonal antibodies also inhibit the growth oftypically less well vascularized human colon cancers in athymic mice anddecrease the number of tumors arising from inoculated cells.

Viral expression of a VEGF-binding construct of Flk-1, Flt-1, the mouseKDR receptor homologue, truncated to eliminate the cytoplasmic tyrosinekinase domains but retaining a membrane anchor, virtually abolishes thegrowth of a transplantable glioblastoma in mice presumably by thedominant negative mechanism of heterodimer formation with membranespanning endothelial cell VEGF receptors. Embryonic stem cells, whichnormally grow as solid tumors in nude mice, do not produce detectabletumors if both VEGF alleles are knocked out. Taken together, these dataindicate the role of VEGF in the growth of solid tumors. Inhibition ofKDR or Flt-1 is implicated in pathological angiogenesis, and thesereceptors are useful in the treatment of diseases in which angiogenesisis part of the overall pathology, e.g., inflammation, diabetic retinalvascularization, as well as various forms of cancer since tumor growthis known to be dependent on angiogenesis. (Weidner et al., N. Engl. J.Med. 324, pp. 1-8. 1991).

Accordingly, the identification of small compounds which specificallyinhibit, regulate and/or modulate the signal transduction of tyrosinekinases is desirable and is an object of this invention.

SUMMARY OF THE INVENTION

The present invention relates to compounds that are capable ofinhibiting, modulating and/or regulating signal transduction of bothreceptor-type and non-receptor type tyrosine kinases. One embodiment ofthe present invention is illustrated by a compound of Formula I, and thepharmaceutically acceptable salts and stereoisomers thereof:

DETAILED DESCRIPTION OF THE INVENTION

The compounds of this invention are useful in the inhibition of kinasesand are illustrated by a compound of Formula I:

or a pharmaceutically acceptable salt or stereoisomer thereof, wherein:

X is CH or N;

R₁ and R3 are independently selected from the group consisting of:

1) H,

2) C₁₋₁₀ alkyl,

3) C₂₋₁₀ alkenyl,

4) C₂₋₁₀ alkynyl,

5) aryl,

6) halo,

7) OH, and

8) heterocyclyl,

said alkyl, alkenyl, alkynyl, aryl, and beterocyclyl is optionallysubstituted with one to three members selected from R^(a);

R₂ is:

1) H,

2) C₁₋₆ alkyl,

3) aryl,

4) OH,

5) NO₂,

6) NH₂, or

7) halogen;

R₅ is:

1) H,

2) C₁₋₆ alkyl,

3) OH,

4) O—C₁₋₆ alkyl,

5) halo,

6) NH₂, or

7) NO₂;

R₇ and R₈ are independently selected from the group consisting of:

1) H,

2) C₁₋₁₀ alkyl,

3) COR,

4) COOR,

5) aryl, and

6) heterocyclyl,

said alkyl, aryl and heterocyclyl optionally substituted with R₉, orNR₇R₈ are be taken together to form a heterocyclic 5-10 memberedsaturated or unsaturated ring containing, in addition to the nitrogenatom, one to two additional heteroatoms selected from the groupconsisting of N, O and S, said ring optionally substituted with one ortwo substituents selected from R^(a);

R₉ is aryl or heterocyclyl, said aryl and heterocyclyl is optionallysubstituted with from one to three members selected from R^(a);

R₁₀ is:

1) H,

2) C₁₋₆ alkyl,

3) NR₇R_(8,)

4) O—C₁₋₆ alkyl,

5) aryl, or

6) heterocyclyl,

said alkyl, aryl, and heterocyclyl is optionally substituted with one tothree members selected from R^(a);

R^(a) is:

1) C₁₋₁₀ alkyl,

2) halogen,

3) NO₂,

4) OR,

5) NR₇R₈,

6) CN,

7) aryl, or

8) heterocyclyl; and

R is H or C₁₋₆ alkyl.

A second embodiment is a compound of Formula I, as described above,wherein

R₁ and R₃ are independently selected from the group consisting of:

1) H,

2) C₁₋₁₀ alkyl,

3) aryl, and

4) heterocyclyl,

said alkyl, aryl, and heterocyclyl is optionally substituted with one tothree members selected from R^(a);

R₂ is:

1) H,

2) C₁₋₆ alkyl,

3) OH, or

4) halogen; and

R₁₀ is:

1) C₁₋₆ alkyl,

2) NR₇R₈,

3) O—C₁₋₆ alkyl,

4) aryl,

5) heterocyclyl,

said aryl and heterocyclyl is optionally substituted with from one tothree members selected from R^(a).

And yet another embodiment is the compound of Formula I wherein X is CH.A further embodiment is a compound as described in the second embodimentabove wherein X is further defined as CH.

And still another embodiment is a compound as described in the secondembodiment above wherein X is further defined as CH and

R₁ is selected from the group consisting of:

1) H,

2) C₁₋₁₀ alkyl,

3) phenyl, and

4) heterocyclyl,

said alkyl phenyl, and heterocyclyl is optionally substituted with oneto three members selected from R^(a);

R₂ is H or C₁₋₆ alkyl;

R₃ is H or C₁₋₃ alkyl;

R₁₀ is:

1) C₁₋₆ alkyl,

2) NR₇R₈,

3) O—C₁₋₆ alkyl,

4) phenyl,

5) heterocyclyl,

said phenyl and heterocyclyl is optionally substituted with from one tothree members selected from R^(a); and

R^(a) is:

1) C₁₋₆ alkyl,

2) halogen,

3) NO₂,

4) OR,

5) NR₇R₈,

6) CN,

7) phenyl, or

8) heterocyclyl.

Yet another embodiment of the present invention is a compound which

is4-(3-phenyl-pyrazolo[1,5a]pyrimidin-6-yl)-1-(3-piperidin-1-yl-propyl)-1H-pyridin-2-one;

1-(2-morpholin-4-yl-ethyl)-4-(3-phenyl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;

1-(3-dimethylamino-propyl)-4-(3-phenyl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;

1-(1-methyl-piperidin-3-ylmethyl)-4-(3-phenyl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;

1-[3-(4-methylpiperazin-1-yl)-propyl)]-4-(3-phenyl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;

1-(2-dimethylamino-propyl)-4-(3-phenyl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;

1-(1-dimethylamino-2-methyl-propyl)-4-(3-phenyl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;

1-[2-(4-cyano-piperidin-1-yl-ethyl]-4-(3-phenyl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;

1-(3-piperidin-1-yl-propyl)-4-(3-thiophen-3-yl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;

1-(3-piperidin-1-yl-ethyl)-4-(3-thiophen-3-yl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;

1-(2-morpholin-4-yl-ethyl)-4-(3-thiophen-3-yl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;

1-(3-dimethylamino-propyl)-4-(3-thiophen-3-yl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;

1-(1-methyl-piperidin-3-ylmethyl)-4-(3-thiophen-3-yl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;

1-[3-(4-methylpiperazin-1-yl)-propyl)]-4-(3-thiophen-3-yl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;

1-(2-dimethylamino-propyl)-4-(3-thiophen-3-yl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;

1-(1-dimethylamino-2-methyl-propyl)-4-(3-thiophen-3-yl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;

1-(3-dimethylamino-propyl)-4-(3-thiophen-3-yl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;

1-[2-(4-cyano-piperidin-1-yl-ethyl]-4-(3-thiophen-3-yl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;

4-(3-phenyl-pyrazolo[1,5-a]pyrimidin-6-yl)-1-(3-piperidin-1-yl-propyl)-1H-pyrimidin-2-one;

1-(2-morpholin-4-yl-ethyl)-4-(3-phenyl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyrimidin-2-one;

1-(3-dimethylamino-propyl)-4-(3-phenyl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyrimidin-2-one;

1-(1-methyl-piperidin-3-ylmethyl)-4-(3-phenyl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyrimidin-2-one;

1-[3-(4-methylpiperazin-1-yl)-propyl)]-4-(3-phenyl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyrimidin-2-one;

1-(2-dimethylamino-propyl)-4-(3-phenyl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyrimidin-2-one;

1-(1-dimethylamino-2-methyl-propyl)-4-(3-phenyl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyrimidin-2-one;and

1-[2-(4-cyano-piperidin-1-yl-ethyl]-4-(3-phenyl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyrimidin-2-one;

or a pharmaceutically acceptable salt or stereoisomer thereof.

Also included within the scope of the present invention is apharmaceutical composition which is comprised of a compound of Formula Ias described above and a pharmaceutically acceptable carrier. Thepresent invention also encompasses a method of treating or preventingcancer in a mammal in need of such treatment which is comprised ofadministering to said mammal a therapeutically effective amount of acompound of Formula I. Preferred cancers for treatment are selected fromcancers of the brain, genitourinary tract, lymphatic system, stomach,larynx and lung. Another set of preferred forms of cancer arehistiocytic lymphoma, lung adenocarcinoma, small cell lung cancers,pancreatic cancer, gioblastomas and breast carcinoma.

Also included is a method of treating or preventing a disease in whichangiogenesis is implicated, which is comprised of administering to amammal in need of such treatment a therapeutically effective amount of acompound of Formula I. Such a disease in which angiogenesis isimplicated is ocular diseases such as retinal vascularization, diabeticretinopathy, age-related macular degeneration, and the like.

Also included within the scope of the present invention is a method oftreating or preventing inflammatory diseases which comprisesadministering to a mammal in need of such treatment a therapeuticallyeffective amount of a compound of Formula I. Examples of suchinflammatory diseases are rheumatoid arthritis, psoriasis, contactdermatitis, delayed hypersensitivity reactions, and the like.

Also included is a method of treating or preventing a tyrosinekinase-dependent disease or condition in a mammal which comprisesadministering to a mammalian patient in need of such treatment atherapeutically effective amount of a compound of Formula I. Thetherapeutic amount varies according to the specific disease and isdiscernable to the skilled artisan without undue experimentation.

A method of treating or preventing retinal vascularization which iscomprised of administering to a mammal in need of such treatment atherapeutically effective amount of compound of Formula I is alsoencompassed by the present invention. Methods of treating or preventingocular diseases, such as diabetic retinopathy and age-related maculardegeneration, are also part of the invention. Also included within thescope of the present invention is a method of treating or preventinginflammatory diseases, such as rheumatoid arthritis, psoriasis, contactdermatitis and delayed hypersensitivity reactions as well as treatmentor prevention of bone associated pathologies selected from osteosarcomaosteoarthritis, and rickets.

The invention also contemplates the use of the instantly claimedcompounds in combination with a second compound selected from the groupconsisting of:

1) an estrogen receptor modulator,

2) an androgen receptor modulator,

3) retinoid receptor modulator,

4) a cytotoxic agent,

5) an antiproliferative agent,

6) a prenyl-protein transferase inhibitor,

7) an HMG-CoA reductase inhibitor,

8) an HIV protease inhibitor,

9) a reverse transcriptase inhibitor, and

10) another angiogenesis inhibitor.

Preferred angiogenesis inhibitors are selected from the group consistingof a tyrosine kinase inhibitor, an inhibitor of epidermal-derived growthfactor, an inhibitor of fibroblast-derived growth factor, an inhibitorof platelet derived growth factor, an MMP inhibitor, an integrinblocker, interferon-α, interleukin-12, pentosan polysulfate, acyclooxygenase inhibitor, carboxyamidotriazole, combretastatin A-4,squalamine, 6-O-chloroacetyl-carbonyl)-fumagillol, thalidomide,angiostatin, troponin-1, and an antibody to VEGF. Preferred estrogenreceptor modulators are tamoxifen and raloxifene.

Also included in the scope of the claims is a method of treating cancerwhich comprises administering a therapeutically effective amount of acompound of Formula I in combination with radiation therapy and/or incombination with a compound selected from the group consisting of:

1) an estrogen receptor modulator,

2) an androgen receptor modulator,

3) retinoid receptor modulator,

4) a cytotoxic agent,

5) an antiproliferative agent,

6) a prenyl-protein transferase inhibitor,

7) an HMG-CoA reductase inhibitor,

8) an HIV protease inhibitor,

9) a reverse transcriptase inhibitor, and

10) another angiogenesis inhibitor.

And yet another embodiment of the invention is a method of treatingcancer which comprises administering a therapeutically effective amountof a compound of Formula I in combination with paclitaxel ortrastuzumab.

These and other aspects of the invention will be apparent from theteachings contained herein.

“Tyrosine kinase-dependent diseases or conditions” refers to pathologicconditions that depend on the activity of one or more tyrosine kinases.Tyrosine kinases either directly or indirectly participate in the signaltransduction pathways of a variety of cellular activities includingproliferation, adhesion and migration, and differentiation. Diseasesassociated with tyrosine kinase activities include the proliferation oftumor cells, the pathologic neovascularization that supports solid tumorgrowth, ocular neovascularization (diabetic retinopathy, age-relatedmacular degeneration, and the like) and inflammation (psoriasis,rheumatoid arthritis, and the like).

The compounds of the present invention may have asymmetric centers,chiral axes, and chiral planes (as described in: E. L. Eliel and S. H.Wilen, Stereochemistry of Carbon Compounds, John Wiley & Sons, New York,1994, pages 1119-1190), and occur as racemates, racemic mixtures, and asindividual diastereomers, with all possible isomers and mixturesthereof, including optical isomers, being included in the presentinvention. In addition, the compounds disclosed herein may exist astautomers and both tautomeric forms are intended to be encompassed bythe scope of the invention, even though only one tautomeric structure isdepicted. For example, any claim to compound A below is understood toinclude tautomeric structure B and vice versa, as well as mixturesthereof.

When any variable (e.g. aryl, heterocycle, R¹, R² etc.) occurs more thanone time in any constituent, its definition on each occurrence isindependent at every other occurrence. Also, combinations ofsubstituents and variables are permissible only if such combinationsresult in stable compounds.

It is understood that substituents and substitution patterns on thecompounds of the instant invention can be selected by one of ordinaryskill in the art to provide compounds that are chemically stable andthat can be readily synthesized by techniques known in the art, as wellas those methods set forth below, from readily available startingmaterials.

As used herein, “alkyl” is intended to include both branched,straight-chain, and cyclic saturated aliphatic hydrocarbon groups havingthe specified number of carbon atoms. For example, C₁-C₁₀, as in “C₁-C₁₀alkyl” is defined to include groups having 1, 2, 3, 4, 5, 6, 7, 8, 9 or10 carbons in a linear, branched, or cyclic arrangement. For example,“C₁-C₁₀ alkyl” specifically includes methyl, ethyl, propyl, butyl,pentyl, hexyl, heptyl, octyl, nonyl, decyl, and so on, as well ascycloalkyls such as cycloproyl, cyclobutyl, cyclopentyl, cyclohexyl,tetrahydronaphthalene, methylenecylohexyl, and so on. “Alkoxy”represents an alkyl group of indicated number of carbon atoms attachedthrough an oxygen bridge.

If no number of carbon atoms is specified, the term “alkenyl” refers toa non-aromatic hydrocarbon radical, straight, branched or cyclic,containing from 2 to 10 carbon atoms and at least one carbon to carbondouble bond. Preferably one carbon to carbon double bond is present, andup to four non-aromatic carbon-carbon double bonds may be present. Thus“C₂-C₆ alkenyl” means an alkenyl radical having from 2 to 6 carbonatoms. Alkenyl groups include ethenyl, propenyl, butenyl andcyclohexenyl. As described above with respect to alkyl, the straight,branched or cyclic portion of the alkenyl group may contain double bondsand may be substituted if a substituted alkenyl group is indicated.

The term “alkynyl” refers to a hydrocarbon radical straight, branched orcyclic, containing from 2 to 10 carbon atoms and at least one carbon tocarbon triple bond. Up to three carbon-carbon triple bonds may bepresent. Thus, “C₂-C₆ alkynyl” means an alkynyl radical having from 2 to6 carbon atoms. Alkynyl groups include ethynyl, propynyl and butynyl. Asdescribed above with respect to alkyl), the straight, branched or cyclicportion of the alkynyl group may contain triple bonds and may besubstituted if a substituted alkynyl group is indicated.

As used herein, “aryl” is intended to mean any stable monocyclic orbicyclic carbon ring of up to 7 atoms in each ring, wherein at least onering is aromatic. Examples of such aryl elements include phenyl,naphthyl, tetrahydronaphthyl, indanyl, biphenyl, phenanthryl, anthryl oracenaphthyl. In cases where the aryl substituent is bicyclic and onering is non-aromatic, it is understood that attachment is via thearomatic ring.

The term heteroaryl, as used herein, represents a stable monocyclic orbicyclic ring of up to 7 atoms in each ring, wherein at least one ringis aromatic and contains from 1 to 4 heteroatoms selected from the groupconsisting of O, N and S. Heteroaryl groups within the scope of thisdefinition include but are not limited to: acridinyl, carbazolyl,cinnolinyl, quinoxalinyl, pyrrazolyl, indolyl, benzotriazolyl, furanyl,thienyl, benzothienyl, benzofuranyl, quinolinyl, isoquinolinyl,oxazolyl, isoxazolyl, indolyl, pyrazinyl, pyridazinyl, pyridinyl,pyrimidinyl, pyrrolyl, tetrahydroquinoline. In cases where theheteroaryl substituent is bicyclic and one ring is non-aromatic orcontains no heteroatoms, it is understood that attachment is via thearomatic ring or via the heteroatom containing ring, respectively.

As appreciated by those of skill in the art “halo” or “halogen” as usedherein is intended to include chloro, fluoro, bromo and iodo. The term“heterocycle” or “heterocyclyl” as used herein is intended to mean a 5-to 10-membered aromatic or nonaromatic heterocycle containing from 1 to4 heteroatoms selected from the group consisting of O, N and S, andincludes bicyclic groups. “Heterocyclyl” therefore includes the abovementioned heteroaryls, as well as dihydro and tetrathydro analogsthereof. Further examples of “heterocyclyl” include, but are not limitedto the following: benzoimidazolyl, benzofuranyl, benzofurazanyl,benzopyrazolyl, benzotriazolyl, benzothiophenyl, benzoxazolyl,carbazolyl, carbolinyl, cinnolinyl, furanyl, imidazolyl, indolinyl,indolyl, indolazinyl, indazolyl, isobenzofuranyl, isoindolyl,isoquinolyl, isothiazolyl, isoxazolyl, naphthpyridinyl, oxadiazolyl,oxazolyl, oxazoline, isoxazoline, oxetanyl, pyranyl, pyrazinyl,pyrazolyl, pyridazinyl, pyridopyridinyl, pyridazinyl, pyridyl,pyrimidyl, pyrrolyl, quinazolinyl, quinolyl, quinoxalinyl,tetrahydropyranyl, tetrazolyl, tetrazolopyridyl, thiadiazolyl,thiazolyl, thienyl, triazolyl, azetidinyl, 1,4-dioxanyl,hexahydroazepinyl, piperazinyl, piperidinyl, pyrrolidinyl, morpholinyl,thiomorpholinyl, dihydrobenzoimidazolyl, dihydrobenzofuranyl,dihydrobenzothiophenyl, dihydrobenzoxazolyl, dihydrofuranyl,dihydroimidazolyl, dihydroindolyl, dihydroisooxazolyl,dihydroisothiazolyl, dihydrooxadiazolyl, dihydrooxazolyl,dihydropyrazinyl, dihydropyrazoyl, dihydropyridinyl, dihydropyrimidinyl,dihydropyrrolyl, dihydroquinolinyl, dihydrotetrazolyl,dihydrothiadiazolyl, dihydrothiazolyl, dihydrothienyl, dihydrotriazolyl,dihydroazetidinyl, methylenedioxybenzoyl, tetrahydrofuranyl, andtetrahydrothienyl, and N-oxides thereof.

The pharmaceutically acceptable salts of the compounds of this inventioninclude the conventional non-toxic salts of the compounds of thisinvention as formed, e.g., from non-toxic inorganic or organic acids.For example, such conventional non-toxic salts include those derivedfrom inorganic acids such as hydrochloric, hydrobromic, sulfuric,sulfamic, phosphoric, nitric and the like: and the salts prepared fromorganic acids such as acetic, propionic, succinic, glycolic, stearic,lactic, malic, tartaric, citric, ascorbic, pamoic, maleic,hydroxymaleic, phenylacetic, glutamic, benzoic, salicylic, sulfanilic,2-acetoxy-benzoic, fumaric, toluenesulfonic, methanesulfonic, ethanedisulfonic, oxalic, isethionic, trifluoroacetic and the like.

In certain instances, R⁷ and R⁸ are defined such that they can be takentogether with the nitrogen to which they are attached to form aheterocyclic 5-10 membered saturated or unsaturated ring containing, inaddition to the nitrogen atom, one to two additional heteroatomsselected from the group consisting of N, O and S, said ring optionallysubstituted with one or two substituents selected from R^(a). Examplesof the 5-7 membered ring systems that can thus be formed include, butare not limited to the following:

The pharmaceutically acceptable salts of the compounds of this inventioncan be synthesized from the compounds of this invention which contain abasic or acidic moiety by conventional chemical methods. Generally, thesalts of the basic compounds are prepared either by ion exchangechromatography or by reacting the free base with stoichiometric amountsor with an excess of the desired salt-forming inorganic or organic acidin a suitable solvent or various combinations of solvents. Similarly,the salts of the acidic compounds are formed by reactions with theappropriate inorganic or organic base.

The compounds of this invention may be prepared by employing reactionsas shown in the following schemes, in addition to other standardmanipulations that are known in the literature or exemplified in theexperimental procedures. These schemes, therefore, are not limited bythe compounds listed nor by any particular substituents employed forillustrative purposes. Substituent numbering as shown in the schemes donot necessarily correlate to that used in the claims.

Generally, a method for the preparation of 3,6-diarylpyrazolo(1,5-A)pyrimidines 3 comprises mixing a commercially availablemalondialdehyde compound 1 with a commercially available aminopyrazole 2in an alcohol, such as ethanol, methanol, isopropanol, butanol and thelike, said alcohol containing catalytic quantities of an acid, such asacetic acid.

Scheme 2 illustrates one possible procedure for making 3,6-diarylpyrazolo(1,5-A)pyrimidines 9 when the desired aminopyrazole is notcommercially available. Compound 8 is obtained via the proceduredescribed in Scheme 1 above. Subsequent treatment of 8 with a boronicacid derivative in the presence of a palladium catalyst provides thedesired compound 9.

Scheme 3 ilustrates an alternative method for the preparation of 3,7diarylpyrazolo(1,5-A)pyrimidines 21. The commercially available ketone(15) and nitrile (18) are treated separately withdimethylformamidedi-methyl acetal (16) in refluxing toluene to giveproduce 17 and 19, respectively. Compound 19 is then treated withhydrazine-hydrochloride in refluxing ethanol to yield the aminopyrazole20. Compounds 17 and 20 are then treated with catalytic amounts ofacetic acid in ethanol as described previously giving the desired of 3,7diarylpyrazolo(1,5-A)pyrimidines 21.

UTILITY

The instant compounds are useful as pharmaceutical agents for mammals,especially for humans, in the treatment of tyrosine kinase dependentdiseases. Such diseases include the proliferation of tumor cells, thepathologic neovascularization (or angiogenesis) that supports solidtumor growth, ocular neovascularization (diabetic retinopathy,age-related macular degeneration, and the like) and inflammation(psoriasis, rheumatoid arthritis, and the like).

The compounds of the instant invention may be administered to patientsfor use in the treatment of cancer. The instant compounds inhibit tumorangiogenesis, thereby affecting the growth of tumors (J. Rak et al.Cancer Research, 55:4575-4580, 1995). The anti-angiogenesis propertiesof the instant compounds may also be useful in the treatment of certainforms of blindness related to retinal vascularization.

The disclosed compounds are also useful in the treatment of certainbone-related pathologies, such as osteosarcoma, osteoarthritis, andrickets, also known as oncogenic osteomalacia. (Hasegawa et al.,Skeletal Radiol., 28, pp.41-45, 1999; Gerber et al., Nature Medicine,Vol. 5, No. 6, pp.623-628, June 1999).

The compounds of this invention may be administered to mammals,preferably humans, either alone or, preferably, in combination withpharmaceutically acceptable carriers or diluents, optionally with knownadjuvants, such as alum, in a pharmaceutical composition, according tostandard pharmaceutical practice. The compounds can be administeredorally or parenterally, including the intravenous, intramuscular,intraperitoneal, subcutaneous, rectal and topical routes ofadministration.

For oral use of a chemotherapeutic compound according to this invention,the selected compound may be administered, for example, in the form oftablets or capsules, or as an aqueous solution or suspension. In thecase of tablets for oral use, carriers which are commonly used includelactose and corn starch, and lubricating agents, such as magnesiumstearate, are commonly added. For oral administration in capsule form,useful diluents include lactose and dried corn starch. When aqueoussuspensions are required for oral use, the active ingredient is combinedwith emulsifying and suspending agents. If desired, certain sweeteningand/or flavoring agents may be added. For intramuscular,intraperitoneal, subcutaneous and intravenous use, sterile solutions ofthe active ingredient are usually prepared, and the pH of the solutionsshould be suitably adjusted and buffered. For intravenous use, the totalconcentration of solutes should be controlled in order to render thepreparation isotonic.

The compounds of the instant invention may also be co-administered withother well known therapeutic agents that are selected for theirparticular usefulness against the condition that is being treated. Forexample, the instant compounds may be useful in combination with knownanti-cancer agents. Such known anti-cancer agents include the following:estrogen receptor modulators, androgen receptor modulators, retinoidreceptor modulators, cytotoxic agents, antiproliferative agents,prenyl-protein transferase inhibitors, HMG-CoA reductase inhibitors, HIVprotease inhibitors, reverse transcriptase inhibitors, and otherangiogenesis inhibitors.

“Estrogen receptor modulators” refers to compounds which interfere orinhibit the binding of estrogen to the receptor, regardless ofmechanism. Examples of estrogen receptor modulators include, but are notlimited to, tamoxifen, raloxifene, idoxifene, LY353381, LY117081,toremifene, fulvestrant,4-[7-(2,2-dimethyl-1-oxopropoxy-4-methyl-2-[4-[2-(1-piperidinyl)ethoxy]phenyl]-2H-1-benzopyran-3-yl]-phenyl-2,2-dimethylpropanoate,4,4′-dihydroxybenzophenone-2,4-dinitrophenylhydrazone, and SH646.

“Androgen receptor modulators” refers to compounds which interfere orinhibit the binding of androgens to the receptor, regardless ofmechanism. Examples of androgen receptor modulators include finasterideand other 5α-reductase inhibitors, nilutamide, flutamide,bicalutamideliarozole, and abiraterone acetate.

“Retinoid receptor modulators” refers to compounds which interfere orinhibit the binding of retinoids to the receptor, regardless ofmechanism. Examples of such retinoid receptor modulators includebexarotene, tretinoin, 13-cis-retinoic acid, 9-cis-retinoic acid,α-difluoromethylornithine, ILX23-7553,trans-N-(4′-hydroxyphenyl)retinamide, N-4-carboxyphenyl retinamide,

“Cytotoxic agents” refer to compounds which cause cell death primarilyby interfering directly with the cell's functioning or inhibit orinterfere with cell myosis, including alkylating agents, tumor necrosisfactors, intercalators, microtubulin inhibitors, and topoisomeraseinhibitors.

Examples of cytotoxic agents include, but are not limited to,tirapazimine, sertenef, cachectin, ifosfamide, tasonermin, lonidamine,carboplatin, altretamine, prednimustine, dibromodulcitol, ranimustine,fotemustine, nedaplatin, oxaliplatin, temozolomide, heptaplatin,estramustine, improsulfan tosilate, trofosfamide, nimustine,dibrospidium chloride, pumitepa, lobaplatin, satraplatin, profiromycin,cisplatin, irofulven, dexifosfamide,cis-aminedichloro(2-methylpyridine)platinum, benzylguanine,glufosfamide, GPX100, (trans, trans,trans)-bis-mu-(hexane-1,6-diamine)-mu-[diamine-platinum(II)]bis[diamine(chloro)platinum(II)]tetrachloride, diarizidinylspermine, arsenic trioxide,1-(11-dodecylamino-10-hydroxyundecyl)-3,7-dimethylxanthine, zorubicin,idarubicin, bisantrene, mitoxantrone, pirarubicin, pinafide, vairubicin,amrubicin, antineoplaston,3′-deamino-3′-morpholino-13-deoxo-10-hydroxycarminomycin, annamycin,galarubicin, elinafide, MEN10755, and4-demethoxy-3-deamino-3-aziridinyl-4-methylsulphonyl-daunorubicin.

Examples of microtubulin inhibitors include paclitaxel, vindesinesulfate, 3′,4′-didehydro-4′-deoxy-8′-norvincaleukoblastine, docetaxol,rhizoxin, dolastatin, mivobulin isethionate, auristatin, cemadotin,RPR109881, BMS184476, vinflunine, ciyptophycin,2,3,4,5,6-pentafluoro-N-(3-fluoro-4-methoxyphenyl)benzene sulfonamide,anhydrovinblastine,N,N-dimethyl-L-valyl-L-valyl-N-methyl-L-valyl-L-prolyl-L-proline-t-butylamide,TDX258, and BMS 188797.

Some examples of topoisomerase inhibitors are topotecan, hycaptamine,irinotecan, rubitecan,6-ethoxypropionyl-3′,4′-O-exo-benzylidene-chartreusin,9-methoxy-N,N-dimethyl-5-nitropyrazolo[3,4,5-k1]acridine-2-(6H)propanamine,1-amino-9-ethyl-5-fluoro-2,3-dihydro-9-hydroxy-4-methyl-1H,12H-benzo[de]pyrano[3′,4′:b,7]indolizino[1,2b]quinoline-10,13(9H,15H)dione,lurtotecan, 7-[2-(N-isopropylamino)ethyl]-(20S)camptothecin, BNP1350,BNPI1100, BN80915, BN80942, etoposide phosphate, teniposide, sobuzoxane,2′-dimethylamino-2′-deoxy-etoposide, GL331,N-[2-(dimethylamino)ethyl]-9-hydroxy-5,6-dimethyl-6H-pyrido[4,3-b]carbazole-1-carboxamide,asulacrine, (5a, 5aB,8aa,9b)-9-[2-[N-[2-(dimethylamino)ethyl)]-N-methylamino]ethyl]-5-[4-hydroxy-3,5-dimethoxyphenyl]-5,5a,6,8,8a,9-hexohydrofuro(3′,4′:6,7)naphtho(2,3-d)-1,3-dioxol-6-one,2,3-(methylenedioxy)-5-methyl-7-hydroxy-8-methoxybenzo[c]-phenanthridinium,6,9-bis[(2-aminoethyl)amino]benzo[g]isoguinoline-5,10-dione,5-(3-aminopropylamino)-7,10-dihydroxy-2-(2-hydroxyethylaminomethyl)-6H-pyrazolo[4,5,1-de]acridin-6-one,N-[1-[2(diethylamino)ethylamino]-7-methoxy-9-oxo-9H-thioxanthen-4-ylmethyl]formamide,N-(2-(dimethylamino)ethyl)acridine-4-carboxamide,6-[[2-(dimethylamino)ethyl]amino]-3-hydroxy-7H-indeno[2,1-c]quinolin-7-one,and dimesna.

“Antiproliferative agents” includes antisense RNA and DNAoligonucleotides such as G3139, ODN698, RVASKRAS, GEM231, and INX3001,and antimetabolites such as enocitabine, carmofur, tegafur, pentostatin,doxifluridine, trimetrexate, fludarabine, capecitabine, galocitabine,cytarabine ocfosfate, fosteabine sodium hydrate, raltitrexed,paltitrexid, emitefur, tiazofurin, decitabine, nolatrexed, pemetrexed,nelzarabine, 2′-deoxy-2′-methylidenecytidine,2′-fluoromethylene-2′-deoxycytidine,N-[5-(2,3-dihydro-benzofuryl)sulfonyl]-N′-(3,4-dichlorophenyl)urea,N6-[4-deoxy-4-[N2-[2(E),4(E)-tetradecadienoyl]glycylamino]-L-glycero-B-L-mannoheptopyranosyl]adenine,aplidine, ecteinascidin, troxacitabine,4-[2-amino-4-oxo-4,6,7,8-tetrahydro-3H-pyrimidino[5,4-b][1,4]thiazin-6-yl-(S)-ethyl]-2,5-thienoyl-L-glutamicacid, aminopterin, 5-flurouracil, alanosine,11-acetyl-8-(carbamoyloxymethyl)-4-formyl-6-methoxy-14-oxa-1,11-diazatetracyclo(7.4.1.0.0)-tetradeca-2,4,6-trien-9-ylacetic acid ester, swainsonine, lometrexol, dexrazoxane, methioninase,2′-cyano-2′-deoxy-N4-palmitoyl-1-B-D-arabino furanosyl cytosine, and3-aminopyridine-2-carboxaldehyde thiosemicarbazone. “Antiproliferativeagents” also includes monoclonal antibodies to growth factors, otherthan those listed under “angiogenesis inhibitors”, such as trastuzumab.

“HMG-CoA reductase inhibitors” refers to inhibitors of3-hydroxy-3-methylglutaryl-CoA reductase. Compounds which haveinhibitory activity for HMG-CoA reductase can be readily identified byusing assays well-known in the art. For example, see the assaysdescribed or cited in U.S. Pat. No. 4,231,938 at col. 6, and WO 84/02131at pp. 30-33. The terms “HMG-CoA reductase inhibitor” and “inhibitor ofHMG-CoA reductase” have the same meaning when used herein.

Examples of HMG-CoA reductase inhibitors that may be used include butare not limited to lovastatin (MEVACOR®; see U.S. Pat. Nos. 4,231,938;4,294,926; 4,319,039), simvastatin (ZOCOR®; see U.S. Pat. Nos.4,444,784; 4,820,850; 4,916,239), pravastatin (PRAVACHOL®; see U.S. Pat.Nos. 4,346,227; 4,537,859; 4,410,629; 5,030,447 and 5,180,589),fluvastatin (LESCOL®; see U.S. Pat. Nos. 5,354,772; 4,911,165;4,929,437; 5,189,164; 5,118,853; 5,290,946; 5,356,896), atorvastatin(LIPITOR®; see U.S. Pat. Nos. 5,273,995; 4,681,893; 5,489,691;5,342,952) and cerivastatin (also known as rivastatin and BAYCHOL®; seeU.S. Pat. No. 5,177,080). The structural formulas of these andadditional HMG-CoA reductase inhibitors that may be used in the instantmethods are described at page 87 of M. Yalpani, “Cholesterol LoweringDrugs”, Chemistry & Industry, pp. 85-89 (Feb. 5, 1996) and U.S. Pat.Nos. 4,782,084 and 4,885,314. The term HMG-CoA reductase inhibitor asused herein includes all pharmaceutically acceptable lactone andopen-acid forms (i.e., where the lactone ring is opened to form the freeacid) as well as salt and ester forms of compounds which have HMG-CoAreductase inhibitory activity, and therefor the use of such salts,esters, open-acid and lactone forms is included within the scope of thisinvention. An illustration of the lactone portion and its correspondingopen-acid form is shown below as structures I and II.

In HMG-CoA reductase inhibitors where an open-acid form can exist, saltand ester forms may preferably be formed from the open-acid, and allsuch forms are included within the meaning of the term “HMG-CoAreductase inhibitor” as used herein. Preferably, the HMG-CoA reductaseinhibitor is selected from lovastatin and simvastatin, and mostpreferably simvastatin. Herein, the term “pharmaceutically acceptablesalts” with respect to the HMG-COA reductase inhibitor shall meannon-toxic salts of the compounds employed in this invention which aregenerally prepared by reacting the free acid with a suitable organic orinorganic base, particularly those formed from cations such as sodium,potassium, aluminum, calcium, lithium, magnesium, zinc andtetramethylammonium, as well as those salts formed from amines such asammonia, ethylenediamine, N-methylglucamine, lysine, arginine,ornithine, choline, N,N′-dibenzylethylenediamine, chloroprocaine,diethanolamine, procaine, N-benzylphenethylamine,1-p-chlorobenzyl-2-pyrrolidine-1′-yl-methylbenzimidazole, diethylamine,piperazine, and tris(hydroxymethyl) aminomethane. Further examples ofsalt forms of HMG-CoA reductase inhibitors may include, but are notlimited to, acetate, benzenesulfonate, benzoate, bicarbonate, bisulfate,bitartrate, borate, bromide, calcium edetate, camsylate, carbonate,chloride, clavulanate, citrate, dihydrochloride, edetate, edisylate,estolate, esylate, fumarate, gluceptate, gluconate, glutamate,glycollylarsanilate, hexylresorcinate, hydrabamine, hydrobromide,hydrochloride, hydroxynapthoate, iodide, isothionate, lactate,lactobionate, laurate, malate, maleate, mandelate, mesylate,methylsulfate, mucate, napsylate, nitrate, oleate, oxalate, pamaote,palmitate, panthothenate, phosphate/diphosphate, polygalacturonate,salicylate, stearate, subacetate, succinate, tannate, tartrate,teoclate, tosylate, triethiodide, and valerate.

Ester derivatives of the described HMG-CoA reductase inhibitor compoundsmay act as prodrugs which, when absorbed into the bloodstream of awarm-blooded animal, may cleave in such a manner as to release the drugform and permit the drug to afford improved therapeutic efficacy.

“Prenyl-protein transferase inhibitor” refers to a compound whichinhibits any one or any combination of the prenyl-protein transferaseenzymes, including farnesyl-protein transferase (FPTase),geranylgeranyl-protein transferase type I (GGPTase-I), andgeranylgeranyl-protein transferase type-II (GGPTase-II, also called RabGGPTase). Examples of prenyl-protein transferase inhibiting compoundsinclude(±)-6-[amino(4-chlorophenyl)(1-methyl-1H-imidazol-5-yl)methyl]-4-(3-chlorophenyl)-1-methyl-2(1H)-quinolinone,(−)-6-[amino(4-chlorophenyl)(1-methyl-1H-imidazol-5-yl)methyl]-4-(3-chlorophenyl)-1-methyl-2(1H)-quinolinone,(+)-6-[amino(4-chlorophenyl)(1-methyl-1H-imidazol-5-yl)methyl]-4-(3-chlorophenyl)-1-methyl-2(1H)-quinolinone,5(S)-n-butyl-1-(2,3-dimethylphenyl)-4-[1-(4-cyanobenzyl)-5-imidazolylmethyl]-2-piperazinone,(S)-1-(3-chlorophenyl)-4-[1-(4-cyanobenzyl)-5-imidazolylmethyl]-5-[2-(ethanesulfonyl)methyl)-2-piperazinone,5(S)-n-Butyl-1-(2-methylphenyl)-4-[1-(4-cyanobenzyl)-5-imidazolylmethyl]-2-piperazinone,1-(3-chlorophenyl)-4-[1-(4-cyanobenzyl)-2-methyl-5-imidazolylmethyl]-2-piperazinone,1-(2,2-diphenylethyl)-3-[N-(1-(4-cyanobenzyl)-1H-imidazol-5-ylethyl)carbamoyl]piperidine,4-{5-[4-Hydroxymethyl-4-(4-chloropyridin-2-ylmethyl)-piperidine-1-ylmethyl]-2-methylimidazol-1-ylmethyl}benzonitrile,4-{5-[4-hydroxymethyl-4-(3-chlorobenzyl)-piperidine-1-ylmethyl]-2-methylimidazol-1-ylmethyl}benzonitrile,4-{3-[4-(2-oxo-2H-pyridin-1-yl)benzyl]-3H-imidazol-4-ylmethyl}benzonitrile,4-{3-[4-(5-chloro-2-oxo-2H-[1,2′]bipyridin-5′-ylmethyl]-3H-imidazol-4-ylmethyl}benzonitrile,4-{3-[4-(2-Oxo-2H-[1,2′]bipyridin-5′-ylmethyl]-3H-imidazol-4-ylmethyl}benzonitrile,4-[3-(2-Oxo-1-phenyl-1,2-dihydropyridin-4-ylmethyl)-3H-imidazol-4-ylmethyl]benzonitrile,18,19-dihydro-19-oxo-5H,17H-6.10:12,16-dimetheno-1H-imidazo[4,3-c][1,11,4]dioxaazacyclo-nonadecine-9-carbonitrile,(±)-19,20-Dihydro-19-oxo-5H-18,21-ethano-12,14-etheno-6,10-metheno-22H-benzo[d]imidazo[4,3-k][1,6,9,12]oxatriaza-cyclooctadecine-9-carbonitrile,19,20-dihydro-19-oxo-5H,17H-18,21-ethano-6,10:12,16-dimetheno-22H-imidazo[3,4-h][1,8,11,14]oxatriazacycloeicosine-9-carbonitrile,and(±)-19,20-Dihydro-3-methyl-19-oxo-5H-18,21-ethano-12,14-etheno-6,10-metheno-22H-benzo[d]imidazo[4,3-k][1,6,9,12]oxa-triazacyclooctadecine-9-carbonitrile.

Other examples of prenyl-protein transferase inhibitors can be found inthe following publications and patents: WO 96/30343, WO 97/18813, WO97/21701, WO 97/23478, WO 97/38665, WO 98/28980, WO 98/29119, WO95/32987, U.S. Pat. No. 5,420,245, U.S. Pat. No. 5,523,430, U.S. Pat.No. 5,532,359, U.S. Pat. No. 5,510,510, U.S. Pat. No. 5,589,485, U.S.Pat. No. 5,602,098, European Pat. Publ. 0 618 221, European Pat. Publ. 0675 112, European Pat. Publ. 0 604 181, European Pat. Publ. 0 696 593,WO 94/19357, WO 95/08542, WO 95/11917, WO 95/12612, WO 95/12572, WO95/10514, U.S. Pat. No. 5,661,152, WO 95/10515, WO 95/10516, WO95/24612, WO 95/34535, WO 95/25086, WO 96/05529, WO 96/06138, WO96/06193, WO 96/16443, WO 96/21701, WO 96/21456, WO 96/22278, WO96/24611, WO 96/24612, WO 96/05168, WO 96/05169, WO 96/00736, U.S. Pat.No. 5,571,792, WO 96/17861, WO 96/33159, WO 96/34850, WO 96/34851, WO96/30017, WO 96/30018, WO 96/30362, WO 96/30363, WO 96/31111, WO96/31477, WO 96/31478, WO 96/31501, WO 97/00252, WO 97/03047, WO97/03050, WO 97/04785, WO 97/02920, WO 97/17070, WO 97/23478, WO97/26246, WO 97/30053, WO 97/44350, WO 98/02436, and U.S. Pat. No.5,532,359. For an example of the role of a prenyl-protein transferaseinhibitor on angiogenesis see European J. of Cancer, Vol. 35, No. 9,pp.1394-1401 (1999).

Examples of HIV protease inhibitors include amprenavir, abacavir,CGP-73547, CGP-61755, DMP450, indinavir, nelfinavir, tipranavir,ritonavir, saquinavir, ABT-378, AG 1776, and BMS-232,632. Examples ofreverse transcriptase inhibitors include delaviridine, efavirenz,GS-840, HB Y097, lamivudine, nevirapine, AZT, 3TC, ddC, and ddI.

“Angiogenesis inhibitors” refers to compounds that inhibit the formationof new blood vessels, regardless of mechanism. Examples of angiogenesisinhibitors include, but are not limited to, tyrosine kinase inhibitors,such as inhibitors of the tyrosine kinase receptors Flt-1 (VEGFR1) andFlk-1/KDR (VEGFR₂₀), inhibitors of epidermal-derived,fibroblast-derived, or platelet derived growth factors, MMP inhibitors,integrin blockers, interferon-α, interleukin-12, pentosan polysulfate,cyclooxygenase inhibitors, including nonsteroidal anti-inflammatories(NSAIDs) like aspirin and ibuprofen as well as selectivecyclooxygenase-2 inhibitors like celecoxib and rofecoxib (PNAS, Vol. 89,p. 7384 (1992); JNCI, Vol. 69, p. 475 (1982); Arch. Opthalmol., Vol.108, p.573 (1990); Anat. Rec., Vol. 238, p. 68 (1994); FEBS Letters,Vol. 372, p. 83 (1995); Clin, Orthop. Vol. 313, p. 76 (1995); J. Mol.Endocrinol., Vol. 16, p.107 (1996); Jpn. J. Phannacol., Vol. 75, p. 105(1997); Cancer Res., Vol. 57, p. 1625 (1997); Cell, Vol. 93, p. 705(1998); Intl. J. Mol. Med. Vol. 2, p. 715 (1998); J. Biol. Chem., Vol.274, p. 9116 (1999)), carboxyamidotriazole, combretastatin A-4,squalamine, 6-O-chloroacetyl-carbonyl)-fumagillol, thalidomide,angiostatin, troponin-1, and antibodies to VEGF. (see, NatureBiotechnology, Vol. 17, pp.963-968 (October 1999); Kim et al., Nature,362, 841-844 (1993)).

Other examples of angiogenesis inhibitors include, but are not limitedto, endostation, ukrain, ranpirnase, IM862,5-methoxy-4-[2-methyl-3-(3-methyl-2-butenyl)oxiranyl]-1-oxaspiro[2,5]oct-6-yl(chloroacetyl)carbamate,acetyldinanaline,5-amino-1-[[3,5-dichloro-4-(4-chlorobenzoyl)phenyl]methyl]-1H-1,2,3-triazole-4-carboxamide,CM101, squalamine, combretastatin, RP14610, NX31838, sulfatedmannopentaose phosphate,7,7-(carbonyl-bis[imino-N-methyl-4,2-pyrrolocarbonylimino[N-methyl4,2-pyrrole]-carbonylimino]-bis-(1,3-naphthalenedisulfonate), and 3-[(2,4-dimethylpyrrol-5-yl)methylene]-2-indolinone(SU5416).

As used above, “integrin blockers” refers to compounds which selectivelyantagonize, inhibit or counteract binding of a physiological ligand tothe α_(v)β₃ integrin, to compounds which selectively antagonize, inhibitor counteract binding of a physiological ligand to the αvβ5 integrin, tocompounds which antagonize, inhibit or counteract binding of aphysiological ligand to both the α_(vβ) ₃ integrin and the α_(vβ) ₅integrin, and to compounds which antagonize, inhibit or counteract theactivity of the particular integrin(s) expressed on capillaryendothelial cells. The term also refers to antagonists of the αvβ₆,α_(v)β₈, α₁β₁, α₂β₁, α₅β₁, α₆β₁ and α₆β₄ integrins. The term also refersto antagonists of any combination of α_(v)β₃, α_(v)β₅, α_(v)β₆, α_(v)β₈,α₁β₁, α₂β₁, α₅β₁, α₆β₁ and α₆β₄ integrins.

Some specific examples of tyrosine kinase inhibitors includeN-(trifluoromethylphenyl)-5-methylisoxazol-4-carboxamide,3-[(2,4-dimethylpyrrol-5-yl)methylidenyl)indolin-2-one,17-(allylamino)-17-demethoxygeldanamycin,4-(3-chloro-4-fluorophenylamino)-7-methoxy-6-[3-(4-morpholinyl)propoxyl]quinazoline,N-(3-ethynylphenyl)-6,7-bis(2-methoxyethoxy)-4-quinazolinamine,BIBX1382,2,3,9,10,11,12-hexahydro-10-(hydroxymethyl)-10-hydroxy-9-methyl-9,12-epoxy-1H-diindolo[1,2,3-fg:3′,2′,1′-kl]pyrrolo[3,4-i][1,6]benzodiazocin-1-one,SH268, genistein, ST1571, CEP2563,4-(3-chlorophenylamino)-5,6-dimethyl-7H-pyrrolo[2,3-d]pyrimidinemethanesulfonate, 4-(3-bromo-4-hydroxyphenyl)amino-6,7-dimethoxyquinazoline,4-(4′-hydroxyphenyl)amino-6,7-dimethoxyquinazoline, SU6668, ST1571A,N4-chlorophenyl-4-(4-pyridylmethyl)-1-phthalazinamine, and EMD121974.

The instant compounds are also useful, alone or in combination withplatelet fibrinogen receptor (GP IIb/IIIMa) antagonists, such astirofiban, to inhibit metastasis of cancerous cells. Tumor cells canactivate platelets largely via thrombin generation. This activation isassociated with the release of VEGF. The release of VEGF enhancesmetastasis by increasing extravasation at points of adhesion to vascularendothelium (Amirkhosravi, Platelets 10, 285-292, 1999). Therefore, thepresent compounds can serve to inhibit metastasis, alone or incombination with GP IIb/IIIa) antagonists. Examples of other fibrinogenreceptor antagonists include abciximab, eptifibatide, sibrafiban,lamifiban. lotrafiban, cromofiban, and CT50352.

If formulated as a fixed dose, such combination products employ thecompounds of this invention within the dosage range described below andthe other pharmaceutically active agent(s) within its approved dosagerange. Compounds of the instant invention may alternatively be usedsequentially with known pharmaceutically acceptable agent(s) when acombination formulation is inappropriate.

The term “administration” and variants thereof (e.g., “administering” acompound) in reference to a compound of the invention means introducingthe compound or a prodrug of the compound into the system of the animalin need of treatment. When a compound of the invention or prodrugthereof is provided in combination with one or more other active agents(e.g., a cytotoxic agent, etc.), “administration” and its variants areeach understood to include concurrent and sequential introduction of thecompound or prodrug thereof and other agents.

As used herein, the term “composition” is intended to encompass aproduct comprising the specified ingredients in the specified amounts,as well as any product which results, directly or indirectly, fromcombination of the specified ingredients in the specified amounts.

The term “therapeutically effective amount” as used herein means thatamount of active compound or pharmaceutical agent that elicits thebiological or medicinal response in a tissue, system, animal or humanthat is being sought by a researcher, veterinarian, medical doctor orother clinician.

The term “treating cancer” or “treatment of cancer” refers toadministration to a mammal afflicted with a cancerous condition andrefers to an effect that alleviates the cancerous condition by killingthe cancerous cells, but also to an effect that results in theinhibition of growth and/or metastasis of the cancer.

The present invention also encompasses a pharmaceutical compositionuseful in the treatment of cancer, comprising the administration of atherapeutically effective amount of the compounds of this invention,with or without pharmaceutically acceptable carriers or diluents.Suitable compositions of this invention include aqueous solutionscomprising compounds of this invention and pharmacologically acceptablecarriers, e.g. saline, at a pH level, e.g. 7.4. The solutions may beintroduced into a patient's blood-stream by local bolus injection.

When a compound according to this invention is administered into a humansubject, the daily dosage will normally be determined by the prescribingphysician with the dosage generally varying according to the age,weight, and response of the individual patient, as well as the severityof the patient's symptoms.

In one exemplary application, a suitable amount of compound isadministered to a mammal undergoing treatment for cancer. Administrationoccurs in an amount between about 0.1 mg/kg of body weight to about 60mg/kg of body weight per day, preferably of between 0.5 mg/kg of bodyweight to about 40 mg/kg of body weight per day.

ASSAYS

The compounds of the instant invention described in the Examples weretested by the assays described below and were found to have kinaseinhibitory activity. Other assays are known in the literature and couldbe readily performed by those of skill in the art. (see, for example,Dhanabal et al., Cancer Res. 59:189-197; Xin et al., J. Biol. Chem.274:9116-9121; Sheu et al., Anticancer Res. 18:4435-4441; Ausprunk etal., Dev. Biol. 38:237-248; Gimbrone et al., J. Natl. Cancer Inst.52:413-427; Nicosia et al., In Vitro 18:538-549).

VEGF RECEPTOR KINASE ASSAY

VEGF receptor kinase activity is measured by incorporation ofradio-labeled phosphate into polyglutamic acid, tyrosine, 4:1 (pEY)substrate. The phosphorylated pEY product is trapped onto a filtermembrane and the incorporation of radio-labeled phosphate quantified byscintillation counting.

MATERIALS

VEGF Receptor Kinase

The intracellular tyrosine kinase domains of human KDR (Terman, B. I. etal. Oncogene (1991) vol. 6, pp. 1677-1683.) and Flt-1 (Shibuya. M. etal. Oncogene (1990) vol. 5, pp. 519-524) were cloned as glutathioneS-transferase (GST) gene fusion proteins. This was accomplished bycloning the cytoplasmic domain of the KDR kinase as an in frame fusionat the carboxy terminus of the GST gene. Soluble recombinant GST-kinasedomain fusion proteins were expressed in Spodoptera frugiperda (Sf21)insect cells (Invitrogen) using a baculovirus expression vector (pAcG2T,Pharmingen).

Lysis Buffer

50 mM Tris pH 7.4, 0.5 M NaCl, 5 mM DTT, 1 mM EDTA, 0.5% triton X-100,10% glycerol, 10 mg/mL of each leupeptin, pepstatin and aprotinin and 1mM phenylmethylsulfonyl fluoride (all Sigma).

Wash Buffer

50 mM Tris pH 7.4, 0.5 M NaCl, 5 mM DTT, 1 mM EDTA, 0.05% triton X-100,10% glycerol, 10 mg/mL of each leupeptin, pepstatin and aprotinin and 1mM phenylmethylsulfonyl fluoride.

Dialysis Buffer

50 mM Tris pH 7.4, 0.5 M NaCl, 5 mM DTT, 1 mM EDTA, 0.05% triton X-100,50% glycerol, 10 mg/mL of each leupeptin, pepstatin and aprotinin and 1mM phenylmethylsuflonyl fluoride.

10×Reaction Buffer

200 mM Tris, pH 7.4, 1.0 M NaCl, 50 mM MnCl₂. 10 mM DTT and 5 mg/mLbovine serum albumin (Sigma).

Enzyme Dilution Buffer

50 mM Tris, pH 7.4, 0.1 M NaCl, 1 mM DTT, 10% glycerol, 100 mg/mL BSA.

10×Substrate

750 μg/mL poly (glutamic acid, tyrosine; 4:1) (Sigma).

Stop Solution

30% trichloroacetic acid, 0.2 M sodium pyrophosphate (both Fisher).

Wash Solution

15% trichloroacetic acid, 0.2 M sodium pyrophosphate.

Filter Plates

Millipore #MAFC NOB, GF/C glass fiber 96 well plate.

METHOD A. Protein Purification

1. Sf21 cells were infected with recombinant virus at a multiplicity ofinfection of 5 virus particles/cell and grown at 27° C. for 48 hours.

2. All steps were performed at 4° C. Infected cells were harvested bycentrifugation at 1000×g and lysed at 4° C. for 30 minutes with{fraction (1/10)} volume of lysis buffer followed by centrifugation at100,000×g for 1 hour. The supernatant was then passed over a glutathioneSepharose column (Pharmacia) equilibrated in lysis buffer and washedwith 5 volumes of the same buffer followed by 5 volumes of wash buffer.Recombinant GST-KDR protein was eluted with wash buffer/10 mM reducedglutathione (Sigma) and dialyzed against dialysis buffer.

B. VEGF Receptor Kinase Assay

1. Add 5 μl of inhibitor or control to the assay in 50% DMSO.

2. Add 35 μl of reaction mix containing 5 μl of 10×reaction buffer, 5 μl25 mM ATP/10 μCi [³³P]ATP (Amersham), and 5 μl 10×substrate.

3. Start the reaction by the addition of 10 μl of KDR (25 nM) in enzymedilution buffer.

4. Mix and incubate at room temperature for 15 minutes.

5. Stop by the addition of 50 μl stop solution.

6. Incubate for 15 minutes at 4° C.

7. Transfer a 90 μl aliquot to filter plate.

8. Aspirate and wash 3 times with wash solution.

9. Add 30 μl of scintillation cocktail, seal plate and count in a WallacMicrobeta scintillation counter.

Human Umbilical Vein Endothelial Cell Mitogenesis Assay

Expression of VEGF receptors that mediate mitogenic responses to thegrowth factor is largely restricted to vascular endothelial cells. Humanumbilical vein endothelial cells (HUVECs) in culture proliferate inresponse to VEGF treatment and can be used as an assay system toquantify the effects of KDR kinase inhibitors on VEGF stimulation. Inthe assay described, quiescent HUVEC monolayers are treated with vehicleor test compound 2 hours prior to addition of VEGF or basic fibroblastgrowth factor (bFGF). The mitogenic response to VEGF or bFGF isdetermined by measuring the incorporation of [³H]thymidine into cellularDNA.

Materials

HUVECs

HUVECs frozen as primary culture isolates are obtained from CloneticsCorp. Cells are maintained in Endothelial Growth Medium (EGM; Clonetics)and are used for mitogenic assays at passages 3-7.

Culture Plates

NUNCLON 96-well polystyrene tissue culture plates (NUNC #167008).

Assay Medium

Dulbecco's modification of Eagle's medium containing 1 g/mL glucose(low-glucose DMEM; Mediatech) plus 10% (v/v) fetal bovine serum(Clonetics).

Test Compounds

Working stocks of test compounds are diluted serially in 100%dimethylsulfoxide (DMSO) to 400-fold greater than their desired finalconcentrations. Final dilutions to IX concentration are made directlyinto Assay Medium immediately prior to addition to cells.

10×Growth Factors

Solutions of human VEGF₁₆₅ (500 ng/mL; R&D Systems) and bFGF (10 ng/mL;R&D Systems) are prepared in Assay Medium.

10×[³H]Thymidine

[Methyl-³H]Thymidine (20 Ci/mmol; Dupont-NEN) is diluted to 80 uCi/mL inlow-glucose DMEM.

Cell Wash Medium

Hank's balanced salt solution (Mediatech) containing 1 mg/mL bovineserum albumin (Boehringer-Mannheim).

Cell Lysis Solution

1 N NaOH, 2% (w/v) Na₂CO₃.

METHOD

1. HUVEC monolayers maintained in EGM are harvested by trypsinizationand plated at a density of 4000 cells per 100 μL Assay Medium per wellin 96-well plates. Cells are growth-arrested for 24 hours at 37° C. in ahumidified atmosphere containing 5% CO₂.

2. Growth-arrest medium is replaced by 100 μL Assay Medium containingeither vehicle (0.25% [v/v] DMSO) or the desired final concentration oftest compound. All determinations are performed in triplicate. Cells arethen incubated at 37° C./5% CO₂ for 2 hours to allow test compounds toenter cells.

3. After the 2-hour pretreatment period, cells are stimulated byaddition of 10 μL/well of either Assay Medium, 10×VEGF solution or10×bFGF solution. Cells are then incubated at 37° C./5% CO₂.

4. After 24 hours in the presence of growth factors, 10×[³H]Thymidine(10 μL/well) is added.

5. Three days after addition of [³H]thymidine. medium is removed byaspiration, and cells are washed twice with Cell Wash Medium (400μL/well followed by 200 μL/well). The washed, adherent cells are thensolubilized by addition of Cell Lysis Solution (100 μL/well) and warmingto 37° C. for 30 minutes. Cell lysates are transferred to 7-mL glassscintillation vials containing 150 μL of water. Scintillation cocktail(5 mL/vial) is added, and cell-associated radioactivity is determined byliquid scintillation spectroscopy.

Based upon the foregoing assays the compounds of formula I areinhibitors of VEGF and thus are useful for the inhibition ofangiogenesis, such as in the treatment of ocular disease, e.g., diabeticretinopathy and in the treatment of cancers, e.g., solid tumors. Theinstant compounds inhibit VEGF-stimulated mitogenesis of human vascularendothelial cells in culture with IC₅₀ values between 0.01-5.0 μM. Thesecompounds also show selectivity over related tyrosine kinases (e.g.,FGFR1 and the Src family; for relationship between Src kinases and VEGFRkinases, see Eliceiri et al., Molecular Cell, Vol. 4, pp.915-924,December 1999).

EXAMPLES

Examples provided are intended to assist in a further understanding ofthe invention. Particular materials employed, species and conditions areintended to be further illustrative of the invention and not limiting ofthe reasonable scope thereof.

Example 1

3-(3-Thiophenyl)-7-(4-pyridyl)pyrazolo(1,5-A)pyrimidine

A 13×100 mm reaction tube was charged with aminopyrazole (22) (16.5 mg,0.100 mmol) dissolved in 0.500 mL EtOH and vinylogous amide (23) (17.6mg, 0.100 mmol) dissolved in 0.200 mL EtOH. Glacial acetic acid (1 drop)was added and the reaction was heated to 80° C. for 14 h. An additional0.100 mL of glacial acetic acid was added and heating was continued foran additional 6 h. The sample was concentrated to dryness to provide thedesired title compound. Analysis by mass spectrometry showed [M+H]⁺279.2.

Example 2

4-(3-Phenyl-pyrazolo[1,5-a]pyrimidin-6-yl)-1-(3-piperidin-1-yl-propyl)-1H-pyridin-2-one

A solution of 2-(4-pyridyl)malondialdehyde (1.00 g, 6.70 mmol, 1 equiv)and 3-amino-4-phenylpyrazole (1.07 g, 6.72 mmol, 1.00 equiv) in absoluteethanol (50 mL) was heated at reflux for 2 h. The reaction mixture wasallowed to cool to 23° C., and the resulting precipitate was filtered,washed with methanol (100 mL), and air dried to give 5 as a light yellowsolid.

mp=226-228° C. ¹H NMR (400 MHz, CDCl₃) δ 8.95 (d, 1H, J=2.4 Hz), 8.85(d, 1H, J=2.4 Hz), 8.78 (atypical dd, 2H, J=4.6, 1.6 Hz), 8.52 (s, 1H),8.06 (dd, 2H, J=8.0, 1.1 Hz), 7.55 (atypical dd, 2H, J=4.6, 1.6 Hz),7.48 (t, 2H, J=7.9 Hz), 7.32 (td, 1H, J=8.2, 0.9 Hz); anal. calcd forC17H12N4: C, 74.98; H, 4.44; N, 20.58. Found C, 75.10; H, 4.56; N,20.50.

A solution of 5 (300 mg, 1.10 mmol, 1 equiv) and 55% mCPBA (449 mg, 1.43mmol, 1.30 equiv) in dichloromethane (50 mL) was stirred at 23° C. for 5h. The precipitate was filtered, washed with dichloromethane (50 ml) andair dried to give 6 as a bright yellow solid.

¹H NMR (400 MHz, (CD₃)₂SO) δ 9.73 (d, 1H, J=2.3 Hz), 9.14 (d, 1H, J=2.3Hz), 8.86 (s, 1H), 8.37 (d, 2H, J=7.0 Hz), 8.18 (d, 2H, J=7.7 Hz),),8.01 (d, 2H, J=7.3 Hz), 7.47 (t, 2H, J=8.0 Hz), 7.28 (t, 1H, J=7.4 Hz).

A solution of 6 (70 mg, 0.24 mmol, 1 equiv) in acetic anhydride (10 mL)was heated at reflux for 16 h. The reaction mixture was allowed to coolto 23° C., then concentrated. To a solution of the residue in anhydrousmethanol (15 mL) was added potassium carbonate (50 mg, 0.36 mmol, 1.5equiv), and the resulting mixture was stirred at 23° C. for 30 min.Excess potassium carbonate was removed by filtration, and the filtratewas concentrated. The residue was partitioned between water (100 mL) andethyl acetate (2×50 mL). The combined organic layers were dried oversodium sulfate and concentrated to leave intermediate 7 as a brownsolid.

¹H NMR (400 MHz, CDCl₃) δ 9.73 (d 1H, J=3Hz), 9.14 (d, 1H, J=2.3 Hz),8.86 (s, 1H), 8.18 (d, 2H, J=7.7 Hz),), 7.53 (d, 1H, J=6.7 Hz), 7.47 (t,2H, J=7.8 Hz), 7.28 (t, 1H, J=7.6 Hz), 6.92 (d, 1H, J=1.2 Hz), 6.75 (dd,1H, J=6.7, 1.2 Hz).

A mixture of unpurified 7 (50 mg, 0.17 mmol, 1 equiv),1-(3-chloropropyl)piperidine hydrochloride (41 mg, 0.21 mmol, 1.2equiv), cesium carbonate (136 mg, 0.416 mmol, 2.4 equiv), and sodiumiodide (31 mg, 0.21 mmol, 1.2 equiv) in DMF (5 mL) was heated at 50° C.for 16 h. More 1-(3-chloropropyl)piperidine hydrochloride (34 mg, 0.17mmol, 1.0 equiv), cesium carbonate (55 mg, 0.17 mmol, 1.0 equiv), andsodium iodide (25 mg, 0.17 mmol, 1.0 equiv) were added and the resultingmixture was heated at 60° C. for 7 h. At this point, additional1-(3-chloropropyl)piperidine hydrochloride (34 mg, 0.17 mmol, 1.0equiv), cesium carbonate (55 mg, 0.17 mmol, 1.0 equiv), and sodiumiodide (25 mg, 0.17 mmol, 1.0 equiv) were added and heating (60° C.) wascontinued for 48 h. The reaction mixture was then partitioned betweenwater (50 mL) and ethyl acetate (3×50 mL). The combined organic layerswere dried over sodium sulfate and concentrated. The residue waspurified by flash column chromatography (CHCl₃ sat'd with NH₃) to affordcompound 8a as a yellow solid.

¹H NMR (400 MHz, CDCl₃) δ 8.88 (d, 1H, J=2.3 Hz), 8.79 (d, 1H, J=2.3Hz),8.51 (s, 1H), 8.05 (d, 2H, J=7.7 Hz),), 7.56 (d, 1H, J=7.0 Hz), 7.48 (t,2H, J=7.8 Hz), 7.31 (t, 1H, J=7.4 Hz), 6.84 (d, 1H, J=2.0 Hz), 6.39 (dd,1H, J=7.0, 2.1 Hz), 4.07 (t, 2H, J=6.7 Hz), 2.36 (br m, 2H), 2.34 (t,2H, J=6.9 Hz), 2.00 (pentet, 2H, J=6.8 Hz), 1.59 (brm, 6H), 1.45 (br m,2H).

What is claimed is:
 1. A method of treating cancer in a mammal in needof such treatment which is comprised of administering to said mammal atherapeutically effective amount of a compound of the formula I:

or a pharmaceutically acceptable salt or stereoisomer thereof, wherein:X is CH; R₁ and R₃ are independently selected from the group consistingof: 1) H, 2) C₁₋₁₀ alkyl, 3) C₂₋₁₀ alkenyl, 4) C₂₋₁₀ alkynyl, 5) aryl,6) halo, 7) OH, and 8) heterocyclyl, said alkyl, alkenyl, alkynyl, aryl,and heterocyclyl is optionally substituted with one to three membersselected from R^(a); R₂ is: 1) H, 2) C₁₋₆ alkyl, 3) aryl, 4) OH, 5) NO₂,6) NH₂, or 7) halogen; R₅ is: 1) H, 2) C₁₋₆ alkyl, 3) OH, 4) O—C₁₋₆alkyl, 5) halo, 6) NH₂, or 7) NO₂; R₇ and R₈ are independently selectedfrom the group consisting of: 1) H, 2) C₁₋₁₀ alkyl, 3) COR, 4) COOR, 5)aryl, and 6) heterocyclyl, said alkyl, aryl and heterocyclyl optionallysubstituted with R₉, or NR₇R₈ are taken together to form a heterocyclic5-10 membered saturated or unsaturated ring containing, in addition tothe nitrogen atom, one to two additional heteroatoms selected from thegroup consisting of N, O and S, said ring optionally substituted withone or two substituents selected from R^(a); R₉ is aryl or heterocyclyl,said aryl and heterocyclyl is optionally substituted with from one tothree members selected from R^(a); R₁₀ is: 1) H, 2) C₁₋₆ alkyl, 3) aryl,or 4) heterocyclyl, said alkyl, aryl, and heterocyclyl is optionallysubstituted with one to three members selected from R^(a); R^(a) is: 1)C₁₋₁₀ alkyl, 2) halogen, 3) NO₂, 4) OR, 5) NR₇R₈, 6) CN, 7) aryl, or 8)heterocyclyl; and R is H or C₁₋₆ alkyl.
 2. The method according to claim1 wherein: R₁ is selected from the group consisting of: 1) H, 2) C₁₋₁₀alkyl, 3) phenyl, and 4) heterocyclyl, said alkyl, phenyl, andheterocyclyl is optionally substituted with one to three membersselected from R^(a); R₂ is H or C₁₋₆ alkyl; R₃ is H or C₁₋₃ alkyl; R₁₀is: 1) C₁₋₆ alkyl, 2) phenyl, 3) heterocyclyl, said phenyl andheterocyclyl is optionally substituted with from one to three membersselected from R^(a); and R^(a) is: 1) C₁₋₆ alkyl, 2) halogen, 3) NO₂, 4)OR, 5) NR₇R₈, 6) CN, 7) phenyl, or 8) heterocyclyl.
 3. The methodaccording to claim 1 wherein the compound of the Formula I is selectedfrom the group consisting of:4-(3-phenyl-pyrazolo[1,5-a]pyrimidin-6-yl)-1-(3-piperidin-1-yl-propyl)-1H-pyridin-2-one;1-(2-morpholin4-yl-ethyl)-4-(3-phenyl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;1-(3-dimethylamino-propyl)-4-(3-phenyl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;1-(1-methyl-piperidin-3-ylmethyl)-4-(3-phenyl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;1-[3-(4-methylpiperazin-1-yl)-propyl)]-4-(3-phenyl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;1-(2-dimethylamino-propyl)-4-(3-phenyl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;1-(1-dimethylamino-2-methyl-propyl)-4-(3-phenyl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;1-[2-(4-cyano-piperidin-1-yl-ethyl]-4-(3-phenyl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pryidin-2-one;1-(3-piperidin-1-yl-propyl)-4-(3-thiophen-3-yl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;1-(3-piperidin-1-yl-ethyl)-4-(3-thiophen-3-yl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridine-2-one;1-(2-morpholin-4-yl-ethyl)-4-(3-thiophen-3-yl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;1-(3-dimethylamino-propyl)-4-(3-thiophen-3-yl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;1-(1-methyl-piperidin-3-ylmethyl)-4-(3-thiophen-3-yl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;1-[3-(4-methylpiperazin-1-yl)-propyl)]-4-(3-thiophen-3-yl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;1-(2-dimethylamino-propyl)-4-(3-thiophen-3-yl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;1-(1-dimethylamino-2-methyl-propyl)-4-(3-thiophen-3-yl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;1-(3-dimethylamino-propyl)-4-(3-thiophen-3-yl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyridin-2-one;1-[2-(4-cyano-piperidin-1-yl-ethyl]-4-(3-thiophen-3-yl-pyrazolo[1,5-a]pyrimidin-6-yl)-1H-pyrimidin-2-one;or a pharmaceutically acceptable salt or stereoisomer thereof.
 4. Amethod of treating cancer in accordance with claim 1 wherein the canceris selected from cancers of the brain, genitourinary tract, lymphaticsystem, stomach, larynx and lung.
 5. A method of treating cancer inaccordance with claim 1 wherein the cancer is selected from histiocyticlymphoma, lung adenocarcinoma, small cell lung cancers, pancreaticcancer, gioblastomas and breast carcinoma.
 6. A method of treating adisease in which angiogenesis is implicated, which is comprised ofadministering to a mammal in need of such treatment a therapeuticallyeffective amount of a compound of the formula I:

or a pharmaceutically acceptable salt or stereoisomer thereof, wherein:X is CH; R₁ and R₃ are independently selected from the group consistingof: 1) H, 2) C₁₋₁₀ alkyl, 3) C₂₋₁₀ alkenyl, 4) C₂₋₁₀ alkynyl, 5) aryl,6) halo, 7) OH, and 8) heterocyclyl, said alkyl, alkenyl, alkynyl, aryl,and heterocyclyl is optionally substituted with one to three membersselected from R^(a); R₂ is: 1) H, 2) C₁₋₆ alkyl, 3) aryl, 4) OH, 5) NO₂,6) NH₂, or 7) halogen; R₅ is: 1) H, 2) C₁₋₆ alkyl, 3) OH, 4) O—C₁₋₆alkyl, 5) halo, 6) NH₂, or 7) NO₂; R₇ and R₈ are independently selectedfrom the group consisting of: 1) H, 2) C₁₋₁₀ alkyl, 3) COR, 4) COOR, 5)aryl, and 6) heterocyclyl, said alkyl, aryl and heterocyclyl optionallysubstituted with R₉, or NR₇R₈ are taken together to form a heterocyclic5-10 membered saturated or unsaturated ring containing, in addition tothe nitrogen atom, one to two additional heteroatoms selected from thegroup consisting of N, O and S, said ring optionally substituted withone or two substituents selected from R^(a); R₉ is aryl or heterocyclyl,said aryl and heterocyclyl is optionally substituted with from one tothree members selected from R^(a); R₁₀ is: 1) H, 2) C₁₋₆ alkyl, 3) aryl,or 4) heterocyclyl, said alkyl, aryl, and heterocyclyl is optionallysubstituted with one to three members selected from R^(a); R^(a) is: 1)C₁₋₁₀ alkyl, 2) halogen, 3) NO₂, 4) OR, 5) NR₇R₈, 6) CN, 7) aryl, or 8)heterocyclyl; and R is H or C₁₋₆ alkyl.
 7. A method in accordance withclaim 6 wherein the disease is an ocular disease.
 8. The methodaccording to claim 6 wherein the disease is an inflammatory disease. 9.A method according to claim 8 wherein the inflammatory disease isselected from rheumatoid arthritis, psoriasis, contact dermatitis anddelayed hypersensitivity reactions.
 10. A method of treating retinalvascularization which is comprised of administering to a mammal in needof such treatment a therapeutically effective amount of compound of theformula I:

or a pharmaceutically acceptable salt or stereoisomer thereof, wherein:X is CH; R₁ and R₃ are independently selected from the group consistingof: 1) H, 2) C₁₋₁₀ alkyl, 3) C₂₋₁₀ alkenyl, 4) C₂₋₁₀ alkynyl, 5) aryl,6) halo, 7) OH, and 8) heterocyclyl, said alkyl, alkenyl, alkynyl, aryl,and heterocyclyl is optionally substituted with one to three membersselected from R^(a); R₂ is: 1) H, 2) C₁₋₆ alkyl, 3) aryl, 4) OH, 5) NO₂,6) NH₂, or 7) halogen; R₅ is: 1) H, 2) C₁₋₆ alkyl, 3) OH, 4) O—C₁₋₆alkyl, 5) halo, 6) NH₂, or 7) NO₂; R₇ and R₈ are independently selectedfrom the group consisting of: 1) H, 2) C₁₋₁₀ alkyl, 3) COR, 4) COOR, 5)aryl, and 6) heterocyclyl, said alkyl, aryl and heterocyclyl optionallysubstituted with R₉, or NR₇R₈ are taken together to form a heterocyclic5-10 membered saturated or unsaturated ring containing, in addition tothe nitrogen atom, one to two additional heteroatoms selected from thegroup consisting of N, O and S, said ring optionally substituted withone or two substituents selected from R^(a); R₉ is aryl or heterocyclyl,said aryl and heterocyclyl is optionally substituted with from one tothree members selected from R^(a); R₁₀ is: 1) H, 2) C₁₋₆ alkyl, 3) aryl,or 4) heterocyclyl, said alkyl, aryl, and heterocyclyl is optionallysubstituted with one to three members selected from R^(a); R^(a) is: 1)C₁₋₁₀ alkyl, 2) halogen, 3) NO₂, 4) OR, 5) NR₇R₈, 6) CN, 7) aryl, or 8)heterocyclyl; and R is H or C₁₋₆ alkyl.
 11. A method of treatingdiabetic retinopathy in a mammal in need of such treatment which iscomprised of administering to said mammal a therapeutically effectiveamount of a compound of the formula I:

or a pharmaceutically acceptable salt or stereoisomer thereof, wherein:X is CH; R₁ and R₃ are independently selected from the group consistingof: 1) H, 2) C₁₋₁₀ alkyl, 3) C₂₋₁₀ alkenyl, 4) C₂₋₁₀ alkynyl, 5) aryl,6) halo, 7) OH, and 8) heterocyclyl, said alkyl, alkenyl, alkynyl, aryl,and heterocyclyl is optionally substituted with one to three membersselected from R^(a); R₂ is: 1) H, 2) C₁₋₆ alkyl, 3) aryl, 4) OH, 5) NO₂,6) NH₂, or 7) halogen; R₅ is: 1) H, 2) C₁₋₆ alkyl, 3) OH, 4) O—C₁₋₆alkyl, 5) halo, 6) NH₂, or 7) NO₂; R₇ and R₈ are independently selectedfrom the group consisting of: 1) H, 2) C₁₋₁₀ alkyl, 3) COR, 4) COOR, 5)aryl, and 6) heterocyclyl, said alkyl, aryl and heterocyclyl optionallysubstituted with R₉, or NR₇R₈ are taken together to form a heterocyclic5-10 membered saturated or unsaturated ring containing, in addition tothe nitrogen atom, one to two additional heteroatoms selected from thegroup consisting of N, O and S, said ring optionally substituted withone or two substituents selected from R^(a); R₉ is aryl or heterocyclyl,said aryl and heterocyclyl is optionally substituted with from one tothree members selected from R^(a); R₁₀ is: 1) H, 2) C₁₋₆ alkyl, 3) aryl,or 4) heterocyclyl, said alkyl, aryl, and heterocyclyl is optionallysubstituted with one to three members selected from R^(a); R^(a) is: 1)C₁₋₁₀ alkyl, 2) halogen, 3) NO₂, 4) OR, 5) NR₇R₈, 6) CN, 7) aryl, or 8)heterocyclyl; and R is H or C₁₋₆ alkyl.
 12. A method of treatingage-related macular degeneration in a mammal in need of such treatmentwhich is comprised of administering to said mammal a therapeuticallyeffective amount of a compound of the formula I:

or a pharmaceutically acceptable salt or stereoisomer thereof, wherein:X is CH; R₁ and R₃ are independently selected from the group consistingof: 1) H, 2) C₁₋₁₀ alkyl, 3) C₂₋₁₀ alkenyl, 4) C₂₋₁₀ alkynyl, 5) aryl,6) halo, 7) OH, and 8) heterocyclyl, said alkyl, alkenyl, alkynyl, aryl,and heterocyclyl is optionally substituted with one to three membersselected from R^(a); R₂ is: 1) H, 2) C₁₋₆ alkyl, 3) aryl, 4) OH, 5) NO₂,6) NH₂, or 7) halogen; R₅ is: 1) H, 2) C₁₋₆ alkyl, 3) OH, 4) O—C₁₋₆alkyl, 5) halo, 6) NH₂, or 7) NO₂; R₇ and R₈ are independently selectedfrom the group consisting of: 1) H, 2) C₁₋₁₀ alkyl, 3) COR, 4) COOR, 5)aryl, and 6) heterocyclyl, said alkyl, aryl and heterocyclyl optionallysubstituted with R₉, or NR₇R₈ are taken together to form a heterocyclic5-10 membered saturated or unsaturated ring containing, in addition tothe nitrogen atom, one to two additional heteroatoms selected from thegroup consisting of N, O and S, said ring optionally substituted withone or two substituents selected from R^(a); R₉ is aryl or heterocyclyl,said aryl and heterocyclyl is optionally substituted with from one tothree members selected from R^(a); R₁₀ is: 1) H, 2) C₁₋₆ alkyl, 3) aryl,or 4) heterocyclyl, said alkyl, aryl, and heterocyclyl is optionallysubstituted with one to three members selected from R^(a); R^(a) is: 1)C₁₋₁₀ alkyl, 2) halogen, 3) NO₂, 4) OR, 5) NR₇R₈, 6) CN, 7) aryl, or 8)heterocyclyl; and R is H or C₁₋₆ alkyl.
 13. A method of treating bonerelated pathologies selected from osteosarcoma, osteoarthritis, andrickets in a mammal in need of such treatment which is comprised ofadministering to said mammal a therapeutically effective amount of acompound of the formula I:

or a pharmaceutically acceptable salt or stereoisomer thereof, wherein:X is CH; R₁ and R₃ are independently selected from the group consistingof: 1) H, 2) C₁₋₁₀ alkyl, 3) C₂₋₁₀ alkenyl, 4) C₂₋₁₀ alkynyl, 5) aryl,6) halo, 7) OH, and 8) heterocyclyl, said alkyl, alkenyl, alkynyl, aryl,and heterocyclyl is optionally substituted with one to three membersselected from R^(a); R₂ is: 1) H, 2) C₁₋₆ alkyl, 3) aryl, 4) OH, 5) NO₂,6) NH₂, or 7) halogen; R5 is: 1) H, 2) C₁₋₆ alkyl, 3) OH, 4) O—C₁₋₆alkyl, 5) halo, 6) NH₂, or 7) NO₂; R₇ and R₈ are independently selectedfrom the group consisting of: 1) H, 2) C₁₋₁₀ alkyl, 3) COR, 4) COOR, 5)aryl, and 6) heterocyclyl, said alkyl, aryl and heterocyclyl optionallysubstituted with R₉, or NR₇R₈ are taken together to form a heterocyclic5-10 membered saturated or unsaturated ring containing, in addition tothe nitrogen atom, one to two additional heteroatoms selected from thegroup consisting of N, O and S, said ring optionally substituted withone or two substituents selected from R^(a); R₉ is aryl or heterocyclyl,said aryl and heterocyclyl is optionally substituted with from one tothree members selected from R^(a); R₁₀ is: 1) H, 2) C₁₋₆ alkyl, 3) aryl,or 4) heterocyclyl, said alkyl, aryl, and heterocyclyl is optionallysubstituted with one to three members selected from R^(a); R^(a) is: 1)C₁₋₁₀ alkyl, 2) halogen, 3) NO₂, 4) OR, 5) NR₇R₈, 6) CN, 7) aryl, or 8)heterocyclyl; and R is H or C₁₋₆ alkyl.